We are proposing a research project that has two aims. We are interested in defining the molecular mechanism of cell fate decision mediated by one of the major eukaryotic phosphatase systems, PP2A/B56. In mammalian systems, PP2A/B56 interacts with major developmental and potential tumor promoting pathways such as Wnt. PP2A/B56 also has been implicated in tumorigenesis and metastasis. Consequently, significant effort has been made to unravel the mechanism of PP2A/B56 mediated cellular events such as development and tumorigenesis. Our proposal is designed to facilitate this quest. One of the complications in studying the role of mammalian PP2A/B56 is the presence of at least five distinct B56 genes and numerous splicing variants, which makes it hard to understand their definite function due to their potential functional redundancy. In contrast, Dictyostelium discoideum, a facultative multicellular organism, has a single B56 homologue and cells lacking B56 were successfully generated by our group. Using Dictyostelium cells lacking B56, we could definitely define its function during development. Re-introduction of B56 restored wild-type like development. By using the null and rescued Dictyostelium system, we will determine the role of PP2A/B56 in cell fate decision processes. Results from Aim1 will shed new light in understanding basic mechanisms of eukaryotic cell differentiation and related pathological processes. The second Aim of the proposal is to define the role of PP2A/B56 under environmental stresses. Previous studies indicated that one of the well-known tumor suppressors, LKB1, is involved in the cellular stress response as well. In mammalian cells, stresses induce activation of cellular energy sensor AMPK, which plays critical roles in the pathogenesis of diabetes as well as development. It is currently not well defined if B56 regulates LKB1 or AMPK. Similar to mammalian cells, Dictyostelium cells also have LKB1 and AMPK. We will also take advantage of Dictyostelium cells lacking B56 to investigate how B56 affect the critical energy sensor in response to oxidative and osmotic stresses. Results from this Specific Aim will facilitate the understanding of cellular response mechanism under stress condition such as inflammation. Public Health Relevance: This proposal is to determine molecular mechanisms of how a single type of cell becomes diverse types of cells and how these cells respond to environmental stresses using a model organism Dictyostelium. Dictyostelium can live as a single cell amoeba or choose to become multi-cellular organism, and is very amenable to laboratory manipulations. These properties put us in a unique position to address proposed questions effectively. Results from the proposed research will fill the gap in the current knowledge generated using different systems.